Advancing nanomedicine with cross-link functionalized nanoparticles for rapid excretion.
Angewandte Chemie (International ed. in English)
Nanoparticles have been widely investigated for preclinical animal models as imaging, therapeutic or theranostic agent. However, a very limited number of nanoscale materials are approved for human use due to retention and toxicity concerns. Recent years have seen in vivo fluorescence imaging in the long end of the second near infrared window (NIR-IIb, 1,500-1,700 nm), affording deeper tissue penetration and higher imaging clarity owing to reduced light scattering and near-zero autofluorescence. Most NIR-IIb fluorophores are nanoparticle based probes with long retentionin the body. Here, we applied a novel cross-linked coating to functionalize core/shell lead sulfide/cadmium sulfide quantum dots (PbS/CdS QDs) emitting at ~1,600 nm. The coating was comprised of an amphiphilic polymer followed by three crosslinked amphiphilic polymeric layers (branched PEG-linear PAA-branched PEG, P 3 coating), imparting high biocompatibility and > 90% excretion of QDs within 2 weeks of intravenous administration. The P 3 -QDs were utilized for conjugation to an engineered anti-CD8 diabody to afford in vivo molecular imaging of CD8+ cytotoxic T lymphocytes (CTLs) in response to anti-PD-L1 therapy. Two-plex molecular imaging in combination with down-conversion Er nanoparticles was performed for real-time in vivo monitoring of PD-L1+tumor cells and CD8+CTLswith cellular resolution by non-invasive NIR-IIb light sheet microscopy (LSM). In another application, angiogenesis in the tumor microenvironment was imaged with P 3 -QDs conjugated to TRC105, a chimeric monoclonal antibody against CD105. Further, P 3 -QDs afforded imaging of lymph nodes deep in the body with a signal-to-background ratio of up to ~170. Lastly, we show that the P 3 coating on magnetic nanoparticles also afforded rapid excretion in < 2 weeks, establishing generality of the approach. The ability of eliminating various nanoparticles from a body opens up many possibilities of nanomedicine for human use.
View details for DOI 10.1002/anie.202008083
View details for PubMedID 32681553
In vivo molecular imaging for immunotherapy using ultra-bright near-infrared-IIb rare-earth nanoparticles.
The near-infrared-IIb (NIR-IIb) (1,500-1,700nm) window is ideal for deep-tissue optical imaging in mammals, but lacks bright and biocompatible probes. Here, we developed biocompatible cubic-phase (alpha-phase) erbium-based rare-earth nanoparticles (ErNPs) exhibiting bright downconversion luminescence at ~1,600nm for dynamic imaging of cancer immunotherapy in mice. We used ErNPs functionalized with cross-linked hydrophilic polymer layers attached to anti-PD-L1 (programmed cell death-1 ligand-1) antibody for molecular imaging of PD-L1 in a mouse model of colon cancer and achieved tumor-to-normal tissue signal ratios of ~40. The long luminescence lifetime of ErNPs (~4.6ms) enabled simultaneous imaging of ErNPs and lead sulfide quantum dots emitting in the same ~1,600nm window. In vivo NIR-IIb molecular imaging of PD-L1 and CD8 revealed cytotoxic T lymphocytes in the tumor microenvironment in response to immunotherapy, and altered CD8 signals in tumor and spleen due to immune activation. The cross-linked functionalization layer facilitated 90% ErNP excretion within 2weeks without detectable toxicity in mice.
View details for DOI 10.1038/s41587-019-0262-4
View details for PubMedID 31570897
- Light-sheet microscopy in the near-infrared II window NATURE METHODS 2019; 16 (6): 545-+
Light-sheet microscopy in the near-infrared II window.
Non-invasive deep-tissue three-dimensional optical imaging of live mammals with high spatiotemporal resolution is challenging owing to light scattering. We developed near-infrared II (1,000-1,700nm) light-sheet microscopy with excitation and emission of up to approximately 1,320nm and 1,700nm, respectively, for optical sectioning at a penetration depth of approximately 750mum through live tissues without invasive surgery and at a depth of approximately 2mm in glycerol-cleared brain tissues. Near-infrared II light-sheet microscopy in normal and oblique configurations enabled in vivo imaging of live mice through intact tissue, revealing abnormal blood flow and T-cell motion in tumor microcirculation and mapping out programmed-death ligand 1 and programmed cell death protein 1 in tumors with cellular resolution. Three-dimensional imaging through the intact mouse head resolved vascular channels between the skull and brain cortex, and allowed monitoring of recruitment of macrophages and microglia to the traumatic brain injury site.
View details for PubMedID 31086342
- A theranostic agent for cancer therapy and imaging in the second near-infrared window NANO RESEARCH 2019; 12 (2): 273–79
A theranostic agent for cancer therapy and imaging in the second near-infrared window.
2019; 12: 273–79
Theranostic nanoparticles are integrated systems useful for simultaneous diagnosis and imaging guided delivery of therapeutic drugs, with wide ranging potential applications in the clinic. Here we developed a theranostic nanoparticle (~ 24 nm size by dynamic light scattering) p-FE-PTX-FA based on polymeric micelle encapsulating an organic dye (FE) fluorescing in the 1,000-1,700 nm second near-infrared (NIR-II) window and an anti-cancer drug paclitaxel. Folic acid (FA) was conjugated to the nanoparticles to afford specific binding to molecular folate receptors on murine breast cancer 4T1 tumor cells. In vivo, the nanoparticles accumulated in 4T1 tumor through both passive and active targeting effect. Under an 808 nm laser excitation, fluorescence detection above 1,300 nm afforded a large Stokes shift, allowing targeted molecular imaging tumor with high signal to background ratios, reaching a high tumor to normal tissue signal ratio (T/NT) of (20.0 ± 2.3). Further, 4T1 tumors on mice were completed eradicated by paclitaxel released from p-FE-PTA-FA within 20 days of the first injection. Pharmacokinetics and histology studies indicated p-FE-PTX-FA had no obvious toxic side effects to major organs. This represented the first NIR-II theranostic agent developed.
View details for DOI 10.1007/s12274-018-2210-x
View details for PubMedID 31832124
View details for PubMedCentralID PMC6907162
- Developing a Bright NIR-II Fluorophore with Fast Renal Excretion and Its Application in Molecular Imaging of Immune Checkpoint PD-L1 ADVANCED FUNCTIONAL MATERIALS 2018; 28 (50)
- Near-Infrared IIb Fluorescence Imaging of Vascular Regeneration with Dynamic Tissue Perfusion Measurement and High Spatial Resolution ADVANCED FUNCTIONAL MATERIALS 2018; 28 (36)
Bright quantum dots emitting at 1,600 nm in the NIR-IIb window for deep tissue fluorescence imaging.
Proceedings of the National Academy of Sciences of the United States of America
2018; 115 (26): 6590–95
With suppressed photon scattering and diminished autofluorescence, in vivo fluorescence imaging in the 1,500- to 1,700-nm range of the near-IR (NIR) spectrum (NIR-IIb window) can afford high clarity and deep tissue penetration. However, there has been a lack of NIR-IIb fluorescent probes with sufficient brightness and aqueous stability. Here, we present a bright fluorescent probe emitting at 1,600 nm based on core/shell lead sulfide/cadmium sulfide (CdS) quantum dots (CSQDs) synthesized in organic phase. The CdS shell plays a critical role of protecting the lead sulfide (PbS) core from oxidation and retaining its bright fluorescence through the process of amphiphilic polymer coating and transferring to water needed for imparting aqueous stability and compatibility. The resulting CSQDs with a branched PEG outer layer exhibited a long blood circulation half-life of 7 hours and enabled through-skin, real-time imaging of blood flows in mouse vasculatures at an unprecedented 60 frames per second (fps) speed by detecting 1,600-nm fluorescence under 808-nm excitation. It also allowed through-skin in vivo confocal 3D imaging of tumor vasculatures in mice with an imaging depth of 1.2 mm. The PEG-CSQDs accumulated in tumor effectively through the enhanced permeation and retention effect, affording a high tumor-to-normal tissue ratio up to 32 owing to the bright 1,600-nm emission and nearly zero autofluorescence background resulting from a large 800-nm Stoke's shift. The aqueous-compatible CSQDs are excreted through the biliary pathway without causing obvious toxicity effects, suggesting a useful class of 1,600-nm emitting probes for biomedical research.
View details for PubMedID 29891702
Molecular Cancer Imaging in the Second Near-Infrared Window Using a Renal-Excreted NIR-II Fluorophore-Peptide Probe
2018; 30 (22): e1800106
In vivo molecular imaging of tumors targeting a specific cancer cell marker is a promising strategy for cancer diagnosis and imaging guided surgery and therapy. While targeted imaging often relies on antibody-modified probes, peptides can afford targeting probes with small sizes, high penetrating ability, and rapid excretion. Recently, in vivo fluorescence imaging in the second near-infrared window (NIR-II, 1000-1700 nm) shows promise in reaching sub-centimeter depth with microscale resolution. Here, a novel peptide (named CP) conjugated NIR-II fluorescent probe is reported for molecular tumor imaging targeting a tumor stem cell biomarker CD133. The click chemistry derived peptide-dye (CP-IRT dye) probe afforded efficient in vivo tumor targeting in mice with a high tumor-to-normal tissue signal ratio (T/NT > 8). Importantly, the CP-IRT probes are rapidly renal excreted (≈87% excretion within 6 h), in stark contrast to accumulation in the liver for typical antibody-dye probes. Further, with NIR-II emitting CP-IRT probes, urethra of mice can be imaged fluorescently for the first time noninvasively through intact tissue. The NIR-II fluorescent, CD133 targeting imaging probes are potentially useful for human use in the clinic for cancer diagnosis and therapy.
View details for PubMedID 29682821
Single-walled carbon nanotubes target neutrophils and Ly-6C(hi) monocytes and localize to joints in murine models of arthritis
AMER ASSOC IMMUNOLOGISTS. 2018
View details for Web of Science ID 000459977703161
3D NIR-II Molecular Imaging Distinguishes Targeted Organs with High-Performance NIR-II Bioconjugates
2018; 30 (13): e1705799
Greatly reduced scattering in the second near-infrared (NIR-II) region (1000-1700 nm) opens up many new exciting avenues of bioimaging research, yet NIR-II fluorescence imaging is mostly implemented by using nontargeted fluorophores or wide-field imaging setups, limiting the signal-to-background ratio and imaging penetration depth due to poor specific binding and out-of-focus signals. A newly developed high-performance NIR-II bioconjugate enables targeted imaging of a specific organ in the living body with high quality. Combined with a home-built NIR-II confocal set-up, the enhanced imaging technique allows 900 µm-deep 3D organ imaging without tissue clearing techniques. Bioconjugation of two hormones to nonoverlapping NIR-II fluorophores facilitates two-color imaging of different receptors, demonstrating unprecedented multicolor live molecular imaging across the NIR-II window. This deep tissue imaging of specific receptors in live animals allows development of noninvasive molecular imaging of multifarious models of normal and neoplastic organs in vivo, beyond the traditional visible to NIR-I range. The developed NIR-II fluorescence microscopy will become a powerful imaging technique for deep tissue imaging without any physical sectioning or clearing treatment of the tissue.
View details for PubMedID 29446156
View details for PubMedCentralID PMC5931222
A bright organic NIR-II nanofluorophore for three-dimensional imaging into biological tissues
2018; 9: 1171
Fluorescence imaging of biological systems in the second near-infrared (NIR-II, 1000-1700 nm) window has shown promise of high spatial resolution, low background, and deep tissue penetration owing to low autofluorescence and suppressed scattering of long wavelength photons. Here we develop a bright organic nanofluorophore (named p-FE) for high-performance biological imaging in the NIR-II window. The bright NIR-II >1100 nm fluorescence emission from p-FE affords non-invasive in vivo tracking of blood flow in mouse brain vessels. Excitingly, p-FE enables one-photon based, three-dimensional (3D) confocal imaging of vasculatures in fixed mouse brain tissue with a layer-by-layer imaging depth up to ~1.3 mm and sub-10 µm high spatial resolution. We also perform in vivo two-color fluorescence imaging in the NIR-II window by utilizing p-FE as a vasculature imaging agent emitting between 1100 and 1300 nm and single-walled carbon nanotubes (CNTs) emitting above 1500 nm to highlight tumors in mice.
View details for PubMedID 29563581
Donor Engineering for NIR-II Molecular Fluorophores with Enhanced Fluorescent Performance
JOURNAL OF THE AMERICAN CHEMICAL SOCIETY
2018; 140 (5): 1715–24
Organic fluorophores have been widely used for biological imaging in the visible and the first near-infrared windows. However, their application in the second near-infrared window (NIR-II, 1000-1700 nm) is still limited mainly due to low fluorescence quantum yields (QYs). Here, we explore molecular engineering on the donor unit to develop high performance NIR-II fluorophores. The fluorophores are constructed by a shielding unit-donor(s)-acceptor-donor(s)-shielding unit structure. Thiophene is introduced as the second donor connected to the shielding unit, which can increase the conjugation length and red-shift the fluorescence emission. Alkyl thiophene is employed as the first donor connected to the acceptor unit. The bulky and hydrophobic alkyl thiophene donor affords larger distortion of the conjugated backbone and fewer interactions with water molecules compared to other donor units studied before. The molecular fluorophore IR-FTAP with octyl thiophene as the first donor and thiophene as the second donor exhibits fluorescence emission peaked at 1048 nm with a QY of 5.3% in aqueous solutions, one of the highest for molecular NIR-II fluorophore reported so far. Superior temporal and spatial resolutions have been demonstrated with IR-FTAP fluorophore for NIR-II imaging of the blood vessels of a mouse hindlimb.
View details for PubMedID 29337545
Near-Infrared IIb Fluorescence Imaging of Vascular Regeneration with Dynamic Tissue Perfusion Measurement and High Spatial Resolution.
Advanced functional materials
2018; 28 (36)
Real-time optical imaging is a promising approach for visualizing in vivo hemodynamics and vascular structure in mice with experimentally induced peripheral arterial disease (PAD). We report the application of a novel fluorescence-based all-optical imaging approach in the near-infrared IIb (NIR-IIb, 1500-1700 nm emission) window, for imaging hindlimb microvasculature and blood perfusion in a mouse model of PAD. In phantom studies, lead sulfide/cadmium sulfide (PbS/CdS) quantum dots showed better retention of image clarity, in comparison with single-walled nanotube (SWNT) NIR-IIa (1000-1400nm) dye, at varying depths of penetration. When systemically injected to mice, PbS/CdS demonstrated improved clarity of the vasculature, compared to SWNTs, as well as higher spatial resolution than in vivo microscopic computed tomography. In a mouse model of PAD, NIR-IIb imaging of the ischemic hindlimb vasculature showed significant improvement in blood perfusion over the course of 10 days (P<0.05), as well as a significant increase in microvascular density over the first 7 days after induction of PAD. In conclusion, NIR-IIb imaging of PbS/CdS vascular contrast agent is a useful multi-functional imaging approach for high spatial resolution imaging of the microvasculature and quantification of blood perfusion recovery.
View details for DOI 10.1002/adfm.201803417
View details for PubMedID 31327961
View details for PubMedCentralID PMC6640151
Developing a Bright NIR-II Fluorophore with Fast Renal Excretion and Its Application in Molecular Imaging of Immune Checkpoint PD-L1.
Advanced functional materials
2018; 28 (50)
Fluorescence imaging in the second near-infrared (NIR-II) window holds impressive advantages of enhanced penetration depth and improved signal-to-noise ratio. Bright NIR-II fluorophores with renal excretion ability and low tissue accumulation are favorable for in vivo molecular imaging applications as they can render the target-mediated molecular imaging process easily distinguishable. Here, a probe (anti-PD-L1-BGP6) comprising a fluorophore (IR-BGP6) covalently bonded to the programmed cell death ligand-1 monoclonal antibody (PD-L1 mAb) for molecular imaging of immune checkpoint PD-L1 (a targeting site upregulated in various tumors for cancer imaging) in the NIR-II window is reported. Through molecular optimization, the bright NIR-II fluorophore IR-BGP6 with fast renal excretion (≈91% excretion in general through urine within the first 10 h postinjection) is developed. The conjugate anti-PD-L1-BGP6 succeeds in profiling PD-L1 expression and realizes efficient noninvasive molecular imaging in vivo, achieving a tumor to normal tissue (T/NT) signal ratio as high as ≈9.5. Compared with the NIR-II fluorophore with high nonspecific tissue accumulation, IR-BGP6 derived PD-L1 imaging significantly enhances the molecular imaging performance, serving as a strong tool for potentially studying underlying mechanism of immunotherapy. The work provides rationales to design renal-excreted NIR-II fluorophores and illustrate their advantages for in vivo molecular imaging.
View details for DOI 10.1002/adfm.201804956
View details for PubMedID 31832053
View details for PubMedCentralID PMC6907024
A high quantum yield molecule-protein complex fluorophore for near-infrared II imaging
Fluorescence imaging in the second near-infrared window (NIR-II) allows visualization of deep anatomical features with an unprecedented degree of clarity. NIR-II fluorophores draw from a broad spectrum of materials spanning semiconducting nanomaterials to organic molecular dyes, yet unfortunately all water-soluble organic molecules with >1,000 nm emission suffer from low quantum yields that have limited temporal resolution and penetration depth. Here, we report tailoring the supramolecular assemblies of protein complexes with a sulfonated NIR-II organic dye (CH-4T) to produce a brilliant 110-fold increase in fluorescence, resulting in the highest quantum yield molecular fluorophore thus far. The bright molecular complex allowed for the fastest video-rate imaging in the second NIR window with ∼50-fold reduced exposure times at a fast 50 frames-per-second (FPS) capable of resolving mouse cardiac cycles. In addition, we demonstrate that the NIR-II molecular complexes are superior to clinically approved ICG for lymph node imaging deep within the mouse body.
View details for DOI 10.1038/ncomms15269
View details for Web of Science ID 000401626200001
View details for PubMedID 28524850
Molecular imaging of biological systems with a clickable dye in the broad 800-to 1,700-nm near-infrared window
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA
2017; 114 (5): 962-967
Fluorescence imaging multiplicity of biological systems is an area of intense focus, currently limited to fluorescence channels in the visible and first near-infrared (NIR-I; ∼700-900 nm) spectral regions. The development of conjugatable fluorophores with longer wavelength emission is highly desired to afford more targeting channels, reduce background autofluorescence, and achieve deeper tissue imaging depths. We have developed NIR-II (1,000-1,700 nm) molecular imaging agents with a bright NIR-II fluorophore through high-efficiency click chemistry to specific molecular antibodies. Relying on buoyant density differences during density gradient ultracentrifugation separations, highly pure NIR-II fluorophore-antibody conjugates emitting ∼1,100 nm were obtained for use as molecular-specific NIR-II probes. This facilitated 3D staining of ∼170-μm histological brain tissues sections on a home-built confocal microscope, demonstrating multicolor molecular imaging across both the NIR-I and NIR-II windows (800-1,700 nm).
View details for DOI 10.1073/pnas.1617990114
View details for PubMedID 28096386
Rational Design of Molecular Fluorophores for Biological Imaging in the NIR-II Window.
A new design for second near-infrared window (NIR-II) molecular fluorophores based on a shielding unit-donor-acceptor-donor-shielding unit (S-D-A-D-S) structure is reported. With 3,4-ethylenedioxy thiophene as the donor and fluorene as the shielding unit, the best performance fluorophores IR-FE and IR-FEP exhibit an emission quantum yield of 31% in toluene and 2.0% in water, respectively, representing the brightest organic dyes in NIR-II region reported so far.
View details for DOI 10.1002/adma.201605497
View details for PubMedID 28117499
Boosting the down-shifting luminescence of rare-earth nanocrystals for biological imaging beyond 1500 nm.
2017; 8 (1): 737
In vivo fluorescence imaging in the near-infrared region between 1500-1700 nm (NIR-IIb window) affords high spatial resolution, deep-tissue penetration, and diminished auto-fluorescence due to the suppressed scattering of long-wavelength photons and large fluorophore Stokes shifts. However, very few NIR-IIb fluorescent probes exist currently. Here, we report the synthesis of a down-conversion luminescent rare-earth nanocrystal with cerium doping (Er/Ce co-doped NaYbF4 nanocrystal core with an inert NaYF4 shell). Ce doping is found to suppress the up-conversion pathway while boosting down-conversion by ~9-fold to produce bright 1550 nm luminescence under 980 nm excitation. Optimization of the inert shell coating surrounding the core and hydrophilic surface functionalization minimize the luminescence quenching effect by water. The resulting biocompatible, bright 1550 nm emitting nanoparticles enable fast in vivo imaging of blood vasculature in the mouse brain and hindlimb in the NIR-IIb window with short exposure time of 20 ms for rare-earth based probes.Fluorescence imaging in the near-infrared window between 1500-1700 nm (NIR-IIb window) offers superior spatial resolution and tissue penetration depth, but few NIR-IIb probes exist. Here, the authors synthesize rare earth down-converting nanocrystals as promising fluorescent probes for in vivo imaging in this spectral region.
View details for PubMedID 28963467
View details for PubMedCentralID PMC5622117
Traumatic Brain Injury Imaging in the Second Near-Infrared Window with a Molecular Fluorophore.
2016; 28 (32): 6872-6879
Traumatic brain injury (TBI) is a leading cause of death and disability worldwide. A bright, renal-excreted, and biocompatible near-infrared II fluorophore for in vivo imaging of TBI is designed. A transient hypoperfusion in the injured cerebral region, followed by fluorophore leakage, is observed. NIR-II fluorophores can provide noninvasive assessment of TBI.
View details for DOI 10.1002/adma.201600706
View details for PubMedID 27253071